A number of gene disruption strains (loss-of-function mutants) of rice have been produced by the property of rice retrotransposon Tos17 that is activated by culture to undergo transposition. Transposons are mutagenic genes which are ubiquitous in the genomes of animals, yeast, bacteria, and plants. Transposons are classified into two categories according to their transposition mechanism. Transposons of class II undergo transposition in the form of DNA without replication. Examples of class II transposons include Ac/Ds, Spm/dSpm and Mu elements of maize (Zea mays) (Fedoroff, 1989, Cell 56, 181-191; Fedoroff et al., 1983, Cell 35, 235-242; Schiefelbein et al., 1985, Proc. Natl. Acad. Sci. USA 82, 4783-4787), and Tam element of Antirrhinum (Antirrhinum majus) (Bonas et al., 1984, EMBO J. 3, 1015-1019). Class II transposons are widely used for gene isolation by means of transposon tagging. Such a technique utilizes a property of transposons, that is, a transposon transposes within a genome and enters a certain gene and, as a result, such a gene is physiologically and morphologically modified, whereby the phenotype controlled by the gene is changed. If such a phenotype change can be detected, the affected gene may be isolated (Bancroft et al., 1993, The Plant Cell, 5, 631-638; Colasanti et al., 1998, Cell, 93, 593-603; Gray et al., 1997, Cell, 89, 25-31; Keddie et al., 1998, The Plant Cell, 10, 877-887; and Whitham et al., 1994, Cell, 78, 1101-1115).
Transposons of class I are also called retrotransposons. Retrotransposons undergo replicative transposition through RNA as an intermediate. A class I transposon was originally identified and characterized in Drosophila and yeast. A recent study has revealed that retrotransposons are ubiquitous and dominant in plant genomes (Bennetzen, 1996, Trends Microbiolo., 4, 347-353; Voytas, 1996, Science, 274, 737-738). It appears that most retrotransposons are integratable but non-transposable units. Recently, it has been reported that some retrotransposons of such a type are activated under stress conditions, such as injury, pathogen attack, and cell culture (Grandbastien, 1998, Trends in Plant Science, 3, 181-187; Wessler, 1996, Curr. Biol., 6, 959-961; Wessler et al., 1995, Curr. Opin. Genet. Devel., 5, 814-821). For example, such activation under stress conditions was found in retrotransposons of tobacco, Tnt1A and Tto1 (Pouteau et al., 1994, Plant J., 5, 535-542; Takeda et al., 1988, Plant Mol. Biol., 36, 365-376), and rice, Tos17 (Hirochlka et al., 1996, Proc. Natl. Acad. Sci. USA, 93, 7783-7788).
The rice retrotransposon Tos17 is a class I element in plants that has been extensively studied. Tos17 was cloned by RT-PCR using degenerate primers prepared based on a conserved amino acid sequence of the reverse transcriptase domains of Ty1-copia group retro-elements (Hirochika et al., 1992, Mol. Gen. Genet., 233, 209-216). Tos17 has a length of 4.3 kb and has two identical LTRs (long terminal repeats) of 138 bp and a PBS (primer binding site) complementary to the 3′ end of the initiator methionine tRNA (Hirochika et al., 1996, supra). Transcription of Tos17 is strongly activated by tissue culture, and the copy number of Tos17 increases with time in culture. Its initial copy number in Nipponbare (a Japonica variety), which is used as a genome research model, is two. In plants regenerated from tissue culture, its copy number is increased to 5 to 30 (Hirochika et al., 1996, supra). Unlike class II transposons found in yeast and Drosophila, Tos17 undergoes random transposition in chromosome and induces stable mutation. Therefore, Tos17 provides a useful tool in reverse genetics for analyzing the function of a gene in rice (Hirochika, 1997, Plant Mol. Biol. 35, 231-240; K. Shimamoto Ed., 1999, Molecular Biology of Rice, Springer-Verlag, 43-58; Miyao et. al., 2003, Plant Cell 15, 1771-1780).